Conventional flow cytometry is a technique which allows for the analysis, counting, and sorting of microscopic particles into subpopulations based on one or more particle characteristics. Typically, a beam of light of a single wavelength is directed onto a hydrodynamically-focused stream of fluid. A number of detectors are aimed at the point where the stream passes through the light beam: one in line with the light beam (forward scatter or FSC) and one or more perpendicular to it (side scatter or SSC) and one or more fluorescent detectors. Each particle entrained in the stream of fluid passing through the beam scatters the light in some way, and fluorescent substances found (either intrinsic or added) in the particle or attached to the particle may be excited into emitting light at a longer wavelength than the light source. This combination of scattered and fluorescent light is received by the detectors, and, by analyzing fluctuations in brightness at each detector, it is then possible to derive various types of information about the physical and chemical structure of each individual particle. FSC typically correlates with the cell size and SSC typically correlates with inner complexity or morphology of the particle (for example shape of the nucleus, the amount and type of cytoplasmic granules or the membrane roughness, or the like).
In particular, the configuration of the flow cell of conventional flow cytometers support a single fluid stream in which particles align to pass in single file through the light beam for interrogation. Because conventional flow cytometers produce only a single fluid stream, the number of particles which can pass through single file through the flow cytometer to be interrogated by the beam of light and associated optical system, detection or computer system can be limited during a period of time.
Additionally, conventional flow cytometers utilize a sheath fluid to hydrodynamically focus the sample fluid stream entraining particles for presentation to the beam of light. Additionally, hydrodynamic focusing to generate a laminar flow of a sample fluid stream within a sheath fluid stream to focus particles requires use of a flow cell having at least a sheath fluid flow path and sample fluid flow path each of which are reduced in diameter to force the particles to the center of the fluid stream. This approach requires a flow cell of substantially greater constructional and operational complexity than if sufficient centration, alignment and spacing of particles could be achieved within a single fluid stream without utilizing sheath fluid. Additionally, providing sheath fluid in the context of a microfluidic chip necessitates an additional fluidic interface between the sheath fluid source and the microfluidic chip.
Moreover, conventional flow cytometers do not concurrently sample particles whether of the same or different types or populations of particles from a corresponding plurality of sample sources, such as 12, 24, 48, 96, 384, or 1536 particle sources, into a corresponding plurality of sample fluid streams for concurrent analysis or separation into subpopulations. Conventional flow cytometers process sample particles in an asynchronous or non-concurrent basis through a single flow cytometry unit. In part, this may be due to the fact that different samples of particles can vary by a number of factors such as viscosity of sample fluid, concentration of particles, size of particles, motility of particles, velocity at which particles can be carried in a fluid stream, particle characteristics analyzed, relative difference in the particle characteristic(s) being analyzed or differentiated, or the like. As a result, each of a plurality of sample fluid streams entraining a corresponding plurality of different particle types or populations, can yield analyzable or sortable events which may occur asynchronously between a plurality of sample fluid streams, but must be concurrently analyzed and sorted. This does not appear to have been achieved utilizing a conventional unitary flow cytometry system. Rather, different particle samples are typically analyzed and sorted using separate flow cytometers each of which analyzes and sorts the particles in a single fluid stream with each analyzable or sortable particle event processed in independent asynchronous serial fashion.
The invention described herein addresses the problems associated with the simultaneous analysis and sorting of a plurality of different particle types in a corresponding plurality of fluid streams in which analyzable or sortable events occur either synchronously or asynchronously between each of the plurality of fluid streams entraining particles.